PROJECT SUMMARY/ABSTRACT Numerous environmental carcinogens and anti-cancer drugs form bulky base adducts in genomic DNA. However, the precise location of these lesions throughout the human genome is not known, and the factors that affect both damage formation and repair are difficult to study with currently available methodologies. Because the genomic location of damage and repair strongly influence the occurrence of pathological conditions, there is a need for new approaches for mapping damage and repair events across the entire genome. The long-term goal of our research program is to better understand how the nucleotide excision repair system targets the removal of bulky base adducts from DNA. The objective of this particular proposal is to further develop and apply novel tools for mapping carcinogen- and chemotherapy-induced DNA damage formation and repair throughout the genome and to identify the key factors that influence both the induction of DNA damage and the efficiency of damage removal by nucleotide excision repair. Our group?s background and expertise in the areas of DNA repair enzymology and genomics makes us uniquely qualified to address this issue. For this proposal, we will focus on identifying the precise locations of DNA base damage formed by the environmental carcinogens ultraviolet (UV) light and benzo[a]pyrene and by platinum-based cancer chemotherapies. We recently developed unique sequencing technologies that we have termed Damage-seq and XR-seq to provide high-resolution DNA sequence information on the formation and repair, respectively, of damage throughout the entire human genome. The rationale for the proposed research is that the ability to map damage and repair may reveal unexpected links between environmental carcinogens, mutagenesis, and human disease at specific genomic sites and suggest new strategies for diagnosing and treating human cancers. Our basic research on DNA adducts in cancer risk and prevention will be examined in the following four specific aims: 1) Method for Quantitative Mapping of DNA Damage Sites (Damage-seq) across the Whole Human Genome; 2) Method for Quantitative Mapping of Excision Repair (XR-seq) of the Whole Human Genome; 3) Genome-wide Analysis of Adduct Formation and Repair as a Function of Differentiation, Cell Cycle, and Chromatin States; and 4) Genome-wide Analysis of Adduct Formation and Repair in Human Biospecimens. The novel methods that we recently developed and will further optimize for mapping damage formation and repair will be used throughout this work. This proposal is innovative because it provides a new and unparalleled approach for characterizing DNA damage induced by environmental carcinogens and anti-cancer drugs. The proposed research is significant because it is expected to significantly expand our understanding of DNA damage formation and repair in the human genome at an unprecedented level of resolution. Ultimately, this knowledge has the potential to improve the prevention strategies for environmental carcinogenesis and to lead to the development of new tools for diagnosing and treating human cancer.